Biochemical analysis of tumor rejection antigen and its role of oncogenesis.

肿瘤排斥抗原的生化分析及其在肿瘤发生中的作用。

• 批准号: 08670252
• 负责人: UENAKA Akiko
• 金额: $ 1.41万
• 依托单位: Okayama University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1996
• 资助国家: 日本
• 起止时间: 1996 至 1997
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08670252/
• 关键词: murine leukemia; tumor rejection antigen; peptide recognized by CTL; proto-oncogene akt; over expression; プロト癌遺伝子akt; 拒絶抗原; CTL認識ペプチド; 原癌遺伝子akt

The antigen peptides recognized by RL*1 specific CTL were determined by direct sequencing of the fraction separated by reversed-phase high performance liquid chromatography. Molecular analysis revealed the antigen peptide were derived from the 5'-untransrated region of proto-oncogene c-akt by insertion of a murine leukemia virus LTR together with six nucleotides of unknown origin. In this study, we investigated this altered Akt molecule and its role of oncogenesis in RL*1.1. Western blot analysis using plyclonal rabbit anti c-Akt synthetic polypeptide antibodies revealed a protein 59 KD in addition to 56 KD normal c-Akt protein in RL*1 cells. 2. The 59 KD altered Akt molecule (RL-Akt) was expressed at about ten times level to normal c-Akt and was detected in the membrane fraction and in the cytosolic fraction, although the c-Akt was detected only in the cytosolic fraction. 3. RL-akt transgeneic NIH 3T3 cells grew well, forming colonies in soft apr. 4. RL-Akt deletion mutant and RL-Akt weakly expressed mutant were established. 5. To assess the effect of RL-Akt in the growth rate in vivo, these mutants and parental RL*1 cells transplanted in BALB/c mice, respectively, parental RL*1 cells grow rapidlly, but weakly expressed mutant grow slowly, and deletion mutant did more slowly.These observations suggests that the over expression and localization in the membrane of RL-Akt resulted in effective signal transduction of growth factor-PI3 pathway and these participates in malignancy in RL*1 leukemia. The study of the effects of RL-Akt to the host immune system are going now.

RL*1特异性CTL识别的抗原肽通过反相高效液相色谱分离的组分的直接测序来确定。分子生物学分析表明，该抗原肽是由原癌基因c-akt的5 ′-非翻译区插入鼠白血病病毒LTR和6个未知核苷酸而得到的。在这项研究中，我们研究了这种改变的Akt分子及其在RL*1.1肿瘤发生中的作用。使用plyclonal兔抗c-Akt合成多肽抗体的Western印迹分析显示，RL*1细胞中除了56 KD的正常c-Akt蛋白外，还存在59 KD的蛋白。2. 59 KD的改变的Akt分子（RL-Akt）的表达水平约为正常c-Akt的10倍，并在膜部分和胞质部分中检测到，虽然c-Akt仅在胞质部分中检测到。3. RL-akt转基因NIH 3 T3细胞生长良好，在柔软的apr. 4中形成集落。建立了RL-Akt缺失突变体和RL-Akt弱表达突变体。5.为了评估RL-Akt对体内生长速率的影响，将这些突变体和亲本RL*1细胞分别移植到BALB/c小鼠中，亲本RL*1细胞生长迅速，而弱表达突变体生长缓慢，这些观察结果表明RL-Akt的过度表达和定位在细胞膜上导致了生长因子的有效信号转导，PI 3通路参与RL*1白血病的恶性转化。RL-Akt对宿主免疫系统的影响的研究正在进行中。

项目成果

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Hashimoto, C. 等人：“成年仓鼠肝脏纯化生长抑制因子的小鼠对应物 cDNA 的分子克隆和序列分析。”

Yokoi, T., et al.: "Diversity of epitopes recognized by cytotoxic T lymphocytes that are specific for rejection antigen peptide pRL1a presented on BALB/c leukemia RL*1." Int.Immunol.9. 1195-1201 (1997)

Yokoi, T. 等人：“细胞毒性 T 淋巴细胞识别的表位多样性，对 BALB/c 白血病 RL*1 上呈现的排斥抗原肽 pRL1a 具有特异性。”

Yokoi,T.,et al.: "Diversity of epitopes recognized by cytotoxic T lymphocytes that are specific for rejection antigen peptide pRLla presented on BALB/c leukemia RL♂1." Int.Immunol.9. 1195-1201 (1997)

Yokoi, T., et al.：“细胞毒性 T 淋巴细胞识别的表位多样性，这些细胞毒性 T 淋巴细胞对 BALB/c 白血病 RL♂1 上呈递的排斥抗原肽 pRLla 具有特异性。Int.Immunol.9 (1997)。”

Chen, B.-G.et al.: "Inhibition by CsA and FK506 of the in vitro proliferative response of gammadelta T cells on stimulation with anti-TCR delta mAb." Transplant Immunol.4. 158-162 (1996)

Chen, B.-G.et al.：“CsA 和 FK506 对抗 TCR delta mAb 刺激下 γδ T 细胞的体外增殖反应的抑制。”

Chen,B.-G.,et al.: "Inhibition by CsA and FK506 of the in vitro proliferative response of γδT cells on stimulation with anti-TCR δ mAb." Transplant Immunol.4. 158-162 (1996)

Chen, B.-G. 等人：“CsA 和 FK506 对抗 TCR δ mAb 刺激下的 γδT 细胞的体外增殖反应的抑制。4 (1996)”